Project Summary Cellular quiescence plays a key role in adult stem cells by providing a pool of self-renewing cells that can differentiate into specific lineages to replenish cells lost from natural turnover or injury. Quiescence also protects cancer stem cells from many common therapies that target rapidly dividing tumors, and is postulated to be a major contributor to cancer relapse. Thus, understanding the regulatory mechanisms that contribute to the self-renewal, survival, and formation of quiescent cells has important implications for many aspects of human health, including tissue regeneration and cancer therapeutics. In this proposal, we will define these mechanisms using the model organism, Saccharomyces cerevisiae, where stationary phase has emerged as an excellent system to study cellular quiescence. Quiescent yeast cells represent a sub-population of growth arrested, non-dividing or G0 cells that can be isolated from a stationary phase culture in sufficient quantities for genetic, molecular, and genomic analyses. Like quiescent adult stem cells in vertebrates, quiescent yeast cells show extended viability and can re-enter the cell cycle when growth-promoting signals are restored. Moreover, the formation of these cells plays a key role in the regulation of chronological lifespan and cellular longevity. In three specific aims, we will use a combination of molecular, genetic, and genomic approaches to define the reproductive capacity of quiescent cells by following the temporal and spatial program of DNA replication upon re-entry of quiescent cells into the cell cycle (Aim1); the mechanisms involved in the repair of exogenous DNA to preserve genome integrity (Aim 2); and the histone post-translational modifications that play a role in the formation and survival of quiescent cells and thereby regulate chronological lifespan (Aim 3). The proposed studies could provide important paradigms for understanding the mechanisms that contribute to the formation and maintenance of quiescent cells in more complex developmental systems, and the relationship of quiescence to cell longevity.